A polyhydroxyalkanoate (hereinafter referred to briefly as “PHA”) is a biodegradable and thermoplastic polyester which is synthesized and accumulated as an energy storage substance in cells of a variety of microorganisms. A PHA, which is produced by microorganisms using natural organic acids or oils as carbon sources, is completely biodegraded by a microorganism in soil or water to be taken up in the carbon cycle of the natural world. Therefore, a PHA can be said to be an environment-conscious plastic material which hardly causes adverse effects for ecological system. In these years, a synthetic plastic came into a serious social problem in view of environment pollution, waste disposal and oil resource, thus a PHA has attracted attention as an eco-friendly green plastic and its practical applications are longed for.
When a PHA is produced on a commercial scale, there is a case in which microorganisms innately producing a PHA are used, or a case in which a PHA synthetic gene is recombinated into a microorganism or a plant to obtain a transformant, which is to be used as a host for production. In both cases, since a PHA is accumulated in the biomasses, the PHA is to be produced by recovering the PHA-containing biomass, and separating and purifying the PHA from the biomass.
As regarding the separation and purification of a PHA from a biomass, a method is known as the most convenient which comprises extracting a PHA using a PHA-soluble solvent, crystallizing the resultant using a poor solvent, and recovering the PHA as a crystal. For example, there is a method comprising drying a biomass in which a PHA is accumulated, extracting the PHA from the dried biomass using a halogen-containing organic solvent such as chloroform and methylene chloride, and then mixing the extract with a poor solvent such as methanol and hexane to precipitate and recover the PHA (see Japanese Kokai Publication Sho-59-205992). With these extraction solvents, a PHA can be extracted from only a dried biomass, therefore a process for drying the biomass obtained from a culture broth is required. In addition, there is such a problem that a halogen-containing organic solvent in connection with the environmental regulation is used.
Japanese Kokai Publication Hei-02-69187 describes a method for extracting a PHB (a homopolymer of 3-hydroxybutyrate) from a wet biomass using a solvent, but all the solvents used in this publication are specific ones such as propanediol and glycerol formal, and are insufficient for commercial scale application from an economical point of view, etc.
Moreover, Japanese Kohyo Publication Hei-10-504460 discloses an extraction using a solvent having preferable water-miscibility. In this publication, methanol, ethanol and isopropanol are mentioned, but with these solvents, a PHA cannot be extracted unless the biomass is treated under pressurized condition at 100° C. or higher, which is far exceeding the boiling point. There is also concern that a significant molecular weight decrease may occur in the dissolution at a high temperature of 140° C. applied in Example 1 or 2. Furthermore, there is described that a hard and opaque gel is formed by cooling, and then the gel is compressed using a rotation roll. However, the present inventors experienced that when a polymer became a hard and opaque gel, it was no longer possible to brush away the gel from a reaction container, and it became substantially impossible to recover a PHA.
In U.S. Pat. No. 5,942,597, a PHA is recovered mainly from a plant using a solvent. In this patent, the recovery is carried out at a PHA concentration of 1%, and the amount to be used of the solvent becomes huge in such low PHA concentration, thus this method is substantially difficult to be applied on a commercial scale. Moreover, the present inventors also experienced that gelation could not be prevented under the above-mentioned conditions, too.
As described above, when a PHA is extracted and recovered using a solvent, since the gelation in crystallization is severe, the solvent extraction method considered to be substantially convenient cannot be used. Alternatively, in order to prevent gelation, there is only a means to carry out dissolution and crystallization at a low PHA concentration. But in this case, since the recovery of a PHA becomes inefficient, it becomes too costly for commercial application in the actual state. As described above, the gelation of PHA is a serious problem. However, even though it has become one of the major causes for obstructing the practical application of a PHA. However, an effective solution which prevents gelation has still not been found.
Accordingly, the subject of the present invention is to provide a method for preventing a polyhydroxyalkanoate from galation, which is a state substantially incapable of being brushed away, and for obtaining its polymer in a quite easily recoverable state, when the polyhydroxyalkanoate is extracted using a solvent from a polyhydroxyalkanoate-containing biomass and the polyhydroxyalkanoate is crystallized.